Preparing a multiple-reagent solution:

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BornAHorn
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Preparing a multiple-reagent solution:

Post by BornAHorn » Mon Aug 25, 2008 7:38 pm

in a scientific paper when it says a solution was homogenized in "10 mL 100 mM Tricine HCl, 10 mM KCl, 1 mM EDTA, 600 mM sucrose (pH 7.5)"

I'm stuck on where to start if to prepare such a solution. If I abritrarily pick say 5 mL Tricine HCl, 1 mL KCl, 1 mL EDTA, and 3 mL sucrose, I can figure out the dilutions necessary from our stock solutions. Otherwise I'm missing a step.

Thanks

blcr11
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Re: Preparing a multiple-reagent solution:

Post by blcr11 » Mon Aug 25, 2008 9:07 pm

Why do you need to be arbitrary? You can be arbitrary in the sense that you can make up whatever volume of buffer is convenient to make given the stock solutions that are available to you. Let’s say your stock solutions are:

1M Tricine-HCl, pH 7.5
1M KCl
0.5M EDTA
2M sucrose

Want 100 ml? Fine. Take:

10 ml 1M Tricine, pH 7.5
1 ml 1M KCl
0.2ml 0.5M EDTA
30 ml 2M sucrose

adjust pH if necessary and dilute to a final volume of 100 ml.

The same will hold for 10 ml if you really only want to make up 10 ml at a time, you just won’t use as great a volume of stock solutions, and it may be a little more difficult to adjust the pH of such a small volume of solution assuming you need to do so. I find it easier to make up a conveniently larger volume of buffer even if most of it goes to waste. You can take as many 10 ml aliquots as you need of prepared buffer, until you run out and have to make some more.

blcr11
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Re: Preparing a multiple-reagent solution:

Post by blcr11 » Mon Aug 25, 2008 9:38 pm

Just to be clear, the idea is to mix the aliquots of stock together in the same container (beaker, graduated cylinder, whatever is convenient--though at some point you will need to be able to measure the final volume of 100 ml, or whatever final volume you're aiming for). After you mix your 10 ml of 1M buffer with 1 ml 1M KCl, etc. you'll have a mixture of 40+ ml of your comingled stock solutions. Because the sucrose is somewhat viscous, you may want to dilute the mixture some before adjusting the pH. The volume at that point isn't too important so long as you don't have to exceed 100 ml total volume as you adjust the pH--if you need to adjust at all. An intermediate working volume of 70-75 ml would probably do fine. Once you adjust the pH as needed, then you dilute the mixture to a final volume of 100 ml and you're done.

BornAHorn
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Re: Preparing a multiple-reagent solution:

Post by BornAHorn » Tue Aug 26, 2008 3:00 am

Thanks, that clears things up. Appreciate your quick and thoughtful response.

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